PROJECT SUMMARY/ABSTRACT Despite the success of modern anti-retroviral therapy (ART) in limiting HIV replication, HIV infection remains a chronic disease creating a substantial burden for both patients and the healthcare system. Moreover, it has become evident that long-term ART will not eliminate HIV, forcing patients to stay on life-long therapy with all its deleterious side effects. Thus, efforts to eradicate HIV infection, or at least induce a state of viral control and drug-free remission are needed. As the virus integrates into the host genome of long-lived T cell populations, it can persist in a latent state creating the viral reservoir and while HIV-1-specific cellular immune responses can partially control virus replication, e.g. as observed in the small subset of HIV elite controllers that have undetectable viral loads in the absence of ART, they have failed in the majority of HIV infected individuals to control or eliminate the viral reservoir. Therapeutic vaccines have been considered to augment virus-specific immune responses to improve host control of virus replication and reduce the size of the viral reservoir. However, frequent viral escape mutations in the reservoir have challenged this concept and thus a therapeutic vaccine strategy will need to induce T-cell responses that have not already experienced immune selection pressure. One strategy to induce broad cellular immune responses is to utilize bioinformatically optimized HIV- 1 ?mosaic? antigens and preclinical data from the non-human primate model confirm that these vaccines expanded cellular immune breadth and result in a significant reduction of median setpoint plasma simian immunodeficiency virus (SIV) levels following ART interruption. Thus, we hypothesize that HIV-specific CD4 and CD8 T-cell responses in chronically infected ART-suppressed individuals can be optimized via therapeutic vaccination with an ad26/MVA vector based vaccine expressing mosaic Gag/Pol and Env epitopes to 1) target novel and/or subdominant epitopes, with 2) cross-reactive T-cell receptor (TCR) clonotypes which 3) will result in enhanced antiviral T-cell efficacy against reservoir viruses. We therefore propose 1) to fine map T-cell responses pre- and post-vaccination, 2) to apply state-of-the-art sequencing technology to determine changes in TCR-repertoire and function and 3) to determine antiviral CD8 T-cell activity against segments of HIV-1 reservoir viruses. Collectively, in this proposal we aim to systematically determine the ability of a cutting edge vaccine regimen to induce functionally enhanced and anti-reservoir T-cell responses with the goal to inform the rational design of therapeutic vaccines optimized for functional HIV cure strategies.